Pectolytic and Cellulolytic Enzymes of Two Populations of Ditylenchus dipsaci on 'Wando' Pea (Pisum sativum L.)

''Wando'' pea is susceptible to Ditylenchus dipsaci from Raleigh, N. C. (RNC) but resistant to the same species from Waynesville, N. C. (WNC). Homogenates of RNC and WNC were analyzed for pectolytic and cellulolytic enzyme activity; both had high C_x activity with WNC two to three times more active than RNC. Polymethylglacturonase activity was three to five times higher in RNC, but polygalacturonase was up to 100 times higher in WNC. Polygalacturonate-trans-eliminase was not detected although a Ca⁺⁺-stimulated pectin methyl-trans-eliminase was present. Enzyme analyses of healthy and infected pea tissue showed only slight enzyme activity unrelated to that in nematode homogenates. No correlation between enzyme activity and the differing pathogenicities could be detected.     

Disc-electrophoretic Patterns of Enzymes and Soluble Proteins of Ditylenchus dipsaci and D. triformis

Soluble protein, esterase and oxidative enzyme patterns of the Waynesville, North Carolina, (WNC) and Raleigh, North Carolina, (RNC) populations of Ditylenchus dipsaci were compared. Polyacrylamide gel electrophoretic patterns of soluble protein extracts of nematodes of the two populations differed. Esterase and catalase patterns, however, were identical. Peroxidatic activity of the catalase isoenzymes from nematodes of the two populations differed when catechol was used as a cosubstrate. Distinct differences were demonstrated in soluble protein and enzyme patterns between D. dipsaci and D. triiormis.     

A Comparison of Pectinases from Ditylenchus dipsaci and Allium cepa Callus Tissue

Ground and whole Ditylenchus dipsaci maintained on onion callus contain no culturable micro-organisms when tested with five check media. Healthy onion callus does not produce pectolytic enzymes. Pectolytic enzymes are present in infected callus. These enzymes are, however, associated with resident nematodes and not host tissues. These results suggest that D. dipsaci is the actual source of the endo-polygalacturonase and endo-pectinmethyltrans-eliminase extracted from them.     

Pectinases in Aqueous Extracts of Ditylenchus dipsaci

Aqueous extracts of a population of Ditylenchus dipsaci isolated from onion and maintained monoxenically on onion callus contained endo-polygalacturonase (endo-PG) and endo-pectinmethyltranseliminase (endo-PMTE). In viscometric tests pH 4.2 and 4.0 were optimal for degradation of sodium polypectate and pectin N.F., respectively, by endo-PG. Endo-PMTE reduced viscosity of pectin N.F. optimally at pH 8.5 or above. Activity was dependent on CaCl₂. Pectinmethylesterase activity was not detected in water, NaCl, or sucrose extracts of these nematodes. The extracts macerated potato tuber tissue, onion cotyledonary tissue, and strips of onion epidermis from the ventral surface of onion bulb scales at pH 4.2, 5.3, and 6.2. Pectin could not be localized with hydroxylamine-ferric chloride reagent in macerated tissues treated for 24 hr with active extract.     

Seasonal Fluctuations of Soil and Tissue Populations of Ditylenchus dipsaci and Aphelenchoides ritzemabosi in Alfalfa

Population dynamics of A. ritzemabosi and D. dipsaci were studied in two alfalfa fields in Wyoming. Symptomatic stem-bud tissue and root-zone soil from alfalfa plants exhibiting symptoms of D. dipsaci infection were collected at intervals of 3 to 4 weeks. Both nematodes were extracted from stem tissue with the Baermann funnel method and from soil with the sieving and Baermann funnel method. Soil moisture and soil temperature at 5 cm accounted for 64.8% and 61.0%, respectively, of the variability in numbers of both nematodes in soil at the Big Horn field. Also at the Big Horn field, A. ritzemabosi was found in soil on only three of the 14 collection dates, whereas D. dipsaci was found in soil on 12 dates. Aphelenchoides ritzemabosi was found in stem tissue samples on 9 of the 14 sampling dates whereas D. dipsaci was found on all dates. Populations of both nematodes in stem tissue peaked in October, and soil populations of both peaked in January, when soil moisture was greatest. Numbers of D. dipsaci in stem tissue were related to mean air temperature 3 weeks prior to tissue collection, while none of the climatic factors measured were associated with numbers of A. ritzemabosi. At the Dayton field, soil moisture plus soil temperature at 5 cm accounted for 98.2% and 91.4% of the variability in the soil populations of A. ritzemabosi and D. dipsaci, respectively. Aphelenchoides ritzemabosi was extracted from soil at two of the five collection dates, compared to extraction of D. dipsaci at three dates. Aphelenchoides ritzemabosi was collected from stem tissue at six of the seven sampling dates while D. dipsaci was found at all sampling dates. The only environmental factor that was associated with an increase in the numbers of both nematodes in alfalfa stem tissue was total precipitation 1 week prior to sampling, and this occurred only at the Dayton field. Numbers of A. ritzemabosi in stem tissue appeared to be not affected by any of the environmental factors studied, while numbers of D. dipsaci in stem tissue were associated with cumulative monthly precipitation, snow cover at time of sampling, and the mean weekly temperature 3 weeks prior to sampling. Harvesting alfalfa reduced the numbers of A. ritzemabosi at the Big Horn field and both nematodes at the Dayton field.     

Superoxide Dismutases: II. Purification and Quantitative Relationship with Water-soluble Protein in Seedlings

Superoxide dismutase was purified from pea (Pisum sativum L., cv. Wando) seeds and corn (Zea mays L., cv. Michigan 500) seedlings. The purified pea enzyme eluting as a single peak from gel exclusion chromatography columns contained the three electrophoretically distinct bands of superoxide dismutase characterizing the crude extract. The purified corn enzyme eluted as the same peak as the pea enzyme, and contained five of the seven active bands found in the crude extract. The similar molecular weights and the cyanide sensitivities of these bands indicated that they are probably isozymes of a cupro-zinc superoxide dismutase. One of the remaining corn bands was shown to be a peroxidase.     

Occurrence of Ditylenchus weischeri and Not D. dipsaci in Field Pea Harvest Samples and Cirsium arvense in the Canadian Prairies

The stem nematode, a parasite of the herbaceous perennial weed, Cirsium arvense (L.) Scop. and identified as Ditylenchus dipsaci (Kühn) Filipjev, was reported in the Canadian prairies in 1979. Recently, D. weischeri Chizhov parasitizing Cirsium arvense was described in Russia, and it has been shown that this species is not an agricultural pest. In this study, we examined Ditylenchus species found in field pea (Pisum sativum L.) grain harvest samples in 2009 and 2010 and from C. arvense shoots in pea fields in the Saskatchewan, Alberta, and Manitoba provinces. Samples from 538 fields (mainly yellow pea) were provided by 151 growers throughout the main pea-growing area of the Canadian prairies. Of the samples collected, 2% were positive for Ditylenchus. The population density of the nematode ranged between 4 and 1,500 nematodes kg^-1 pea harvest sample and related to presence of C. arvense seeds. Positive samples occurred in 2009 but not in 2010 and were from throughout the pea-growing area of the Canadian prairies and not related to cropping history. C. arvense collected from yellow pea fields in Saskatchewan and Manitoba, but not Alberta, were infested with Ditylenchus. Morphological and molecular (ITS-PCR-RFLP) traits indicated that this species belongs to D. weischeri. The results indicated the stem nematode found in yellow pea grain is D. weischeri which resided with C. arvense seeds and debris to pea samples. Unlike D. dipsaci, D. weischeri is not a nematode pest of economic importance; therefore, its presence in the pea harvest samples was not a concern.     

Nematodes associated with peas and beans in Scotland

Six hundred and sixty-four soil samples from farms growing peas and beans in eastern Scotland were examined for plant parasitic and predatory nematodes. Neither pea cyst-nematode, Heterodera goettingiana nor pea early-browning virus disease were found. Stem nematode, Ditylenchus dipsaci was recorded in broad beans once. The most frequently occurring plant parasitic nematodes were Tylenchorhynchus dubius, Longidorus elongatus, Rotylenchus goodeyi and Helicotylenchus vulgaris but numbers were generally too small to cause damage. It is concluded that although potentially damaging plant parasitic nematodes occur in Scotland, nematode-related diseases occur infrequently and are of little significance to the crop.     

Pectolytic enzyme activity from Nicotiana tabacum pollen

Ungerminated pollen of Nicotiana tabacum contains a pectolytic enzyme which has its optimal activity between pH 5.5 and 6.5. Pectic lyase was not detected.     

The Interrelationship of Heterodera schachtii and Ditylenchus dipsaci on Sugarbeet

Heterodera schachtii significantly (P = 0.05) reduced sugarbeet root growth below that of uninoculated controls at 20, 24, and 28 C, and Ditylenchus dipsaci significantly (P = 0.05) reduced root growth below that of uninoculated controls at 16, 20, 24, and 28 C. A combination of H. schachtii and D. dipsaci significantly (P = 0.05) reduced root growth below that of single inoculations of H. schachtii at all temperatures and D. dipsaci at 20, 24, and 28 C. Single inoculations of H. schachtii and D. dipsaci significantly (P = 0.05) reduced top growth of sugarbeet below that of uninoculated controls at 20, 24, and 28 C, and 16, 20, 24, and 28 C, respectively. A combination of the two nematodes significantly (P = 0.05) reduced top growth below that of single inoculations of H. schachtii at all temperatures. However, a combination of the two nematodes failed to significantly (P = 0.05) reduce top growth below that of single inoculations of D. dipsaci at any temperature. Inoculations of either H. schachtii or D. dipsaci did not affect penetration of the other nematode, and D. dipsaci did not affect development and reproduction of H. schachtii. D. dipsaci did not reproduce on sugarbeet.     

Separation of Three Species of Ditylenchus and Some Host Races of D. dipsaci by Restriction Fragment Length Polymorphism

This study examined the ribosomal cistron of Ditylenchus destructor, D. myceliophagus and seven host races of D. dipsaci from different geographic locations. The three species showed restriction fragment length polymorphisms (RFLPs) in the ribosomal cistron, the 18S rDNA gene, and the ribosomal internal transcribed spacer (ITS). Southern blot analysis with a 7.5-kb ribosomal cistron probe differentiated the five host races of D. dipsaci examined. Polymerase chain reaction (PCR) amplification of the ITS, followed by digestion with some restriction endonucleases (but not others), produced restriction fragments diagnostic of the giant race. Because the PCR product from D. myceliophagus and the host races of D. dipsaci was about 900 base pairs and the ITS size in D. destructor populations was 1,200 base pairs, mixtures of populations could be detected by PCR amplification. ITS fragments differentiated between D. dipsaci and Aphelenchoides rhyntium in mixed populations. This study establishes the feasibility of differentiation of the host races of D. dipsaci by probing Southern blots with the whole ribosomal cistron.     

Interactions Among Selected Endoparasitic Nematodes and Three Pseudomonads on Alfalfa

Meloidogyne hapla, Pratylenchus penetrans, and Helicotylenchus dihystera, reduced the growth of ''Saranac AR alfalfa seedlings when applied at concentrations of 50 nematodes per plant. All except P. penetrans reduced seedling growth when applied at 25 per seedling. M. hapla reduced growth when applied at 12 per seedling. Nematodes interacted with three pseudomonads to produce greater growth reductions than were obtained with single pathogens, suggesting synergistic relationships. Ditylenchus dipsaci, applied at 25 or 50 nematodes per seedling, reduced plant weight compared with weights of control plants, but did not interact with test bacteria. All of the nematodes except D. dipsaci produced root wounds which were invaded by bacteria.     

Sex Expression and Tail Morphology of Female Progenies of Smooth-Tail and Crenate-Tail Females of Pratylenchus penetrans

An analysis of the offspring of single smooth- and crenate-tail females of Pratylenchus penetrans indicated the existence of progenies containing only males or females. Of the 80 progenies analyzed, 46 contained females with smooth and crenate tails. In general, regardless of the mother''s tail type, most females possessed crenate tails, although more crenate-tail females originated from a crenate-tail female than from a smooth-tail female. Twenty-three progenies contained only females with crenate tails, most of them originating from crenate-tail females. One progeny originating from a smooth-tail female contained only females with smooth tails. No simple interpretation of the inheritance of tail type could be attempted because selection pressure favored females with crenate tails when P. penetrans was reared on Wando pea plants.     

Anhydrobiosis in nematodes—I. The role of glycerol myo-inositol and trehalose during desiccation

• 1.1. The concentrations of free glycerol, inositol and trehalose in five species of nematodes were determined. Analyses of total inositol content were also made.
• 2.2. Significant differences in free and bound sugar levels were found between the two good anhydrobiotes Anguina tritici and Ditylenchus dipsaci and the three poor survivors Pangrellus redivivus, D. myceliophagous and Turbatrix aceti.
• 3.3. Highest trehalose contents were found in desiccated A. tritici and D. dipsaci, but glycerol levels were low.
• 4.4. P. redivivus and T. aceti contained high concentrations of free glycerol.
• 5.5. Desiccated A. tritici larvae contained more free and bound inositol than all other species studied, but desiccated D. dipsaci larvae had higher levels of bound inositol than P. redivivus, D. myceliophagous and T. aceti.
• 6.6. Dramatic reductions in inositol and trehalose contents were found in revived A. tritici larvae and freshly extracted D. dipsaci larvae. This was accompanied by an increase in glycerol content.
• 7.7. The results are discussed in relation to the possible biochemical adaptations employed by anhydrobiotes during desiccation.

     

Gall Formation on Cirsium arvense by Ditylenchus dipsaci

Ditylenchus dipsaci was found to cause gall formation on the stems of Cirsium arvense. The galls were characterized by extensive hypertrophy and hyperplasia, differentiation of nutritive tissue, nuclear modification, and a central cavity containing nematodes. These findings emphasize the importance of host response in investigations of host-parasite interactions and suggest that D. dipsaci may be evolving a host race by reproductive isolation within the confines of a plant gall.     

Control of Heterodera carotae,Ditylenchus dipsaci,and Meloidogyne javanica with Fumigant and Nonfumigant Nematicides

Five field trials were conducted in Italy in 1983 and 1984 to test the efficacy of isazofos and benfuracarb in controlling Heterodera carotae on carrot, Ditylenchus dipsaci on onion, and Meloidogyne javanica on tomato. Methyl isothiocyanate (MIT) was tested against H. carotae and M. javanica. Single (10 kg a.i./ha) and split (5 + 5 kg a.i./ha) applications of isazofos gave yield increases of carrot and onion similar to those obtained with DD (300 liters/ha) and aldicarb (10 kg a.i./ha). Population densities of H. carotae in carrot roots at harvest and of M. javanica in tomato roots 2 months after transplanting were also suppressed by isazofos. Benfuracarb (10 kg a.i./ha increased onion yields in a field infested with D. dipsaci, but it was not effective against H. carotae or M. javanica. The efficacy of MIT at 400 and 600 liters/ha was similar to that of MIT + DD (Di-Trapex) at 300 liters/ha. Both nematicides inhibited hatch of H. carotae eggs and decreased the soil population density of M. javanica.     

Managing Nematode Population Densities on Tomato Transplants Using Crop Rotation and a Nematicide

Millet, milo, soybean, crotalaria, and Norman pigeon pea were used in conjunction with clean fallow and a nematicide (fensulfothion) for managing nematode populations in the production of tomato transplants (Lycopersicon esculentum Mill.). Glean fallow was the most effective treatment in suppressing nematode numbers. After 2 years in tomato, root-knot nematodes increased in numbers to damaging levels, and fallow was no longer effective for complete control even in conjunction with fensulfothion. After 4 years in tomato, none of the crops used as summer cover crops alone or in conjunction with fensulfothion reduced numbers of root-knot nematodes in harvested tomato transplants sufficiently to meet Georgia certification regulations. Milo supported large numbers of Macroposthonia ornata and Pratylenchus spp. and crotalaria supported large numbers of Pratylenchus spp. Millet, milo, soybean, crotalaria, and pigeon pea are poor choices for summer cover crops in sites used to produce tomato transplants, because they support large populations of root-knot and other potentially destructive nematodes.     

Parasitism of Nonhost Cultivars by Ditylenchus dipsaci

The alfalfa race of Ditylenchus dipsaci parasitized and caused characteristic symptoms on nonhost seedlings of sweet clover, onion, tomato, sugarbeet, and wheat in controlled growth-chamber studies. Although the nematode was unable to reproduce on any of the cultivars, it caused plant mortality ranging from 20% on sugarbeet and tomato to 100% on onion.     

Ditylenchus dipsaci Infestation of Trifolium repens. II. Dynamics of Infestation Development

Trifolium repens (white clover) stolons were inoculated with Ditylenchus dipsaci (stem nematode), and the development of resulting infestations was monitored. Nematodes initially remained confined to superficial locations, concentrating in petiole axils near inoculation points. They were able to migrate slowly from the inidal inoculation points and infest adjacent axils, especially in regions near the stolon tip. As time progressed, in some axils, nematodes migrated through the stolon epidermis and colonized slowly expanding subepidermal pockets of host tissue (ca. 0.2-mm length of stolon/day). In these loci nematodes established exponentially increasing populations, but the rates of locus expansion remained constant, indicating that locus expansion was limited by unidentified host-dependent factors. As a result of increasing population pressure within subepidermal loci, J4 entered a "diapause" state and the rate of egg production by adults declined, thereby reducing rate of population growth to more sustainable levels. Typically, these populations peaked at ca. 10,000 individuals in ca. 160 days occupying 3-cm lengths of stolon. Thereafter, heavily infested regions of stolons started to die, leading to the formation of longitudinal splits in their epidermis. In other axils, nematodes did not migrate into the stolons but remained confined to axils. Some of these populations increased a hundred-fold in 95 days, with population growth ending when petioles started to die. Host plant stolon morphology was affected only when subepidermal stolon populations developed high population levels (>100 nematodes) within close proximity (<2 cm) to active terminal meristems. This occurred either when axillary buds became active on previously infested nodes or when nematodes established endoparasitic populations at locations near the stolon tip during winter and spring, when the rate of stolon extension was limited by low light intensity. Affected stolon tips could "escape" from the influence of such infestations when light intensity and temperature increased. Nematode activity was limited by low temperature rather than light intensity. Global warming is likely to lead to greater damage to infested plants during the winter and early spring because the predicted milder winter temperatures will enhance nematode activity but not necessarily promote stolon growth.     

Comparative Calorie Values of Nematodes

Calorie values for a wide biological selection of nematodes, determined with a microbomb calorimeter, ranged from 3.86 to 6.85 Kcal/g. The mean of 5.095 Kcal from 16 species was lower than means recorded in three previous studies of other invertebrate groups. The nematode values were skewed to the lowest limit. Larvae of Ditylenchus dipsaci showed lower calorie values after storage, and the calorie values of separate tissues of Ascaris lumbricoides were highest for eggs and the intestine and lowest for cuticle and body-wall musculature. No clear calorie distinction exists between nematodes with a parasitic or free-living habit or between large and small nematodes.